The present invention relates to immunochemical analysis techniques and more particularly to rate nephelometric immunochemical analysis.
In copending United States patent application No. 796,621, by Robert J. Anderson et al. entitled A SYSTEM FOR RATE IMMUNONEPHELOMETRIC ANALYSIS, assigned to the common assignee of this application, a system is described for antigen/antibody analysis employing rate nephelometric techniques. The rate of change with respect to time of a nephelometric signal obtained from an antibody-antigen reaction is monitored on a continuing basis. The maximum or peak rate of change of the reaction and the time to that peak rate is determined. Over a portion of a curve obtained by plotting peak rate (H) v. antigen concentration (C) for the particular reaction the curve is taken as linear so that by limiting the analysis to that falling on the "linear" portion of the curve the concentration is taken to be substantially equal to a constant times the peak rate.
Such a conclusion is, at best, true over only a very limited portion of the curve. Likewise, the amount of the curve which will behave in a linear manner is a function of the particular antigen-antibody reaction being monitored. In actuality, the curve of peak rate (H) v. antigen concentration (C) is appreciably nonlinear and cannot be fit by a conventional power series of less than fourth order, e.g., H=a.sub.0 +a.sub.1 C+a.sub.2 C.sup.2 +a.sub.3 C.sup.3 +a.sub.4 C.sup.4 + . . .
Where a relationship of such complexity is involved, normal practice would be to determine the unknown antigen concentration C by plotting and then referring to a calibration curve H v. C. Such a curve would be generated by measuring peak rates obtained with a set of known dilutions of an antigen sample (calibrator) of known concentration. The set would usually include at least five different dilutions. Having generated the calibration curve, the unknown concentration of an antigen in an antibody-antigen reaction would be determined by measuring the peak rate and then reading the concentration value which corresponds to the measured peak rate on the calibration curve.
The foregoing is a tedious manual procedure. It is, therefore, desirable to obtain an approximation equation for the H v. C curve which will permit the mathematical calculation of unknown concentration values (C) within acceptable limits from measured values of peak rates (H). It is further desirable that the approximation equation be solvable for the concentration values on a real-time basis, as by iterative methods, within a reasonable time constraint, to provide maximum throughput in a clinical situation.
Therefore, it is the object of the present invention to provide a method of determining concentration from peak rate data in a rate nephelometric immunochemical analysis system employing a mathematical approximation of at least the antibody excess portion of the H v. C curve.